mmg_233_2013_genetics_genomicswikiaorg-20200214-history
Antisense Oligonucleotide
Background Huntington's Disease is an autosomal dominant neurodegenerative disorder resulting from trinucleotide expansion within the huntingtin gene (Htt). Afflicted individuals harbor a mutant allele encoding more than 36 CAG repeats within Htt. Onset of symptoms is directly related to the number of these repeats and typically occurs between the ages of 35 and 44 with changes in mood or cognitive ability. Symptoms progress in a predictable manner with the loss of coordination giving rise to uncontrolled tremors and dementia, ultimately leading to death within twenty years of onset 1. The role Huntingtinin within the cell is complex poorly understo od, although it has been implicated in coordinating neuronal development and physiology through its affect as a transcrip tion factor. In addition, Htt has recently been shown to play a role in regulating membrane vesicle dynamics, mitochondrial efficiency, and endosomal recycling 2. Furthermore, the protein has been shown to associate with over 100 cytoplasmic targets within the neuron 3. As the accumulation of mutated Htt is toxic to cells and leads to premature neuron death overtime. Strategy for developing a Htt knockdown therapeautic Although many different treatment options have been explored over the years, none have proven successful at halting disease progression. Recently however, gene silencing strategies utlizing Htt-specfic antisense oligonucleotides and small interfering RNAs have been described with promising in vitro results 4. Several approaches have been explored to deliver antisense Huntingtinin oligonucleotides and siRNAs to target neurons. The most successful strategy involves packaging these molecules into an adenoviral vector before they are administered to the target tissue (the brain) through intracerebroventricular injection 5. Once in the cells, the antisense oligonucleotides and siRNAs are able to bind their target mRNA in the cytoplasm. In the case of siRNA, the doublestranded hybrid triggers ]the RNA silencing machinery, which will then selectively degrade all of the Huntingtinin mRNA within the cell. In the case of the antisense oligonucleotides, the doublestranded hybrid either triggers the degradation of the Huntingtinin mRNA through the activity of RNAse H or physically blocks its translation into protein. These molecules are designed to target either the affected (mutant) Htt allele or transcripts produced by both alleles. Current challanges As with most gene therapy strategies, properly targeting the desired tissue has proven to be a difficult. Achieving consistent, in vivo ''delivery of siRNAs and antisense oligos to the brain has been a major challenge in developing a reliable gene silencing based therapeutic for treating Huntington Disease. Since these molecules are poorly absorbed from the gastrointestinal tract, specifically targeting neurons in the brain via oral delivery is not an option. As a result, siRNAs and antisense oligos require direct delivery to the brain through either intraparenchymal or intracerebroentrivular routes in order to effectively lower Htt levels 5. Another challenge has been to determine the proper dosage needed to sufficiently maintain lower levels of the mutated protein without completely eliminating the expression of the wild type protein. Studies have shown that deletion of ''Htt in mice is lethal within the first 18 days of development, and that complete knockdown within the developed brain leads to premature death. Currently, allele-specific approaches are being explored to target mutant transcripts for degradation while leaving the levels of wild type Htt transcripts unaffected. Preliminary results from an in vitro study have provided some promise, where a sevenfold enrichment for the wild type transcript were observed. References 1 http://en.wikipedia.org/wiki/Huntington%27s_disease 2 Carroll JB, Warby SC, Southwell AL, Doty CN, Greenlee S, Skotte N, Hung G, Bennett CF, Freier SM, Hayden MR. Potent and selective antisense oligonucleotides targeting single-nucleotide polymorphisms in the Huntington disease gene / allele-specific silencing of mutant huntingtin. Mol Ther. 2011;19:2178–2185. 3 Nucifora FC, Jr, et al. Interference by huntingtin and atrophin-1 with cbp-mediated transcription leading to cellular toxicity. Science. 2001;291(5512):2423–2428. doi: 10.1126/science.1056784. 4 DiFiglia M, et al. Therapeutic silencing of mutant huntingtin with siRNA attenuates striatal and cortical neuropathology and behavioral deficits. Proc Natl Acad Sci U S A. 2007;104(43):17204–17209. doi: 10.1073/pnas.0708285104. 5 Dinah W.Y. Sah and Neil Aronin. Oligonucleotide therapeutic approaches for Huntington disease. J Clin Invest. 2011 February 1; 121(2): 500–507.